ETHNOPHARMACOLOGICAL RELEVANCE:Aucubin is one of the major bioactive iridoid glycosides present in Rehmanniae radix preparata, a traditional Chinese medicinal herb that has long been used to nourish the kidney, benefit the brain, and calm the mind, and is widely applied in the treatment of neurological and neuropsychiatric disorders. Modern pharmacological studies have demonstrated that aucubin exerts prominent anti-inflammatory and neuroprotective effects. Aucubin has been shown to ameliorate attention deficit hyperactivity disorder (ADHD)-like behaviors; however, the underlying mechanisms remain largely unknown. AIM OF THE STUDY:This study aimed to investigate the neuroprotective effects of aucubin against ADHD-like behaviors and to elucidate the underlying molecular mechanisms. METHODS:To elucidate the protective effects of Aucubin against ADHD through network pharmacology, molecular docking analysis, and experimental validation. An ADHD-like phenotype in offspring mice was established through intraperitoneal administration of S-ketamine in the middle and late trimesters of gestation. Aucubin (40 mg/kg) was intraperitoneally injected into offspring mice 14 days after birth, once daily for seven consecutive days. To assess behavioral, electrophysiological, and pathological changes in mice, several tests were employed, including the open field test (OFT), novel object recognition (NOR) test, elevated plus-maze (EPM) test, fear conditioning (FC), local field potential recording, and immunofluorescence assays. Furthermore, results in vitro have investigated the impact of aucubin on S-ketamine-induced astrocytic damage. RESULTS:Aucubin administration significantly reversed S-ketamine-induced ADHD-like behaviors, including decreased total distance in the OFT, increased recognition index in the NOR, reduced proportion of EPM's open-arm timing, and increased freezing time in the FC. Additionally, it decreased the glial fibrillary acidic protein intensity and theta (θ) oscillation power during NOR. In vitro studies demonstrated the neuroprotective effects of aucubin. Furthermore, NLRP3 activator Nigericin inhibited the neuroprotective effects of aucubin. CONCLUSIONS:Aucubin significantly improved ADHD-like behavior in mice by inhibiting NLRP3 activity in astrocytes.

BACKGROUND:Diabetic kidney disease (DKD) is a major microvascular complication of diabetes, with microcirculatory dysfunction and immune injury as its core pathological features. Extracellular vesicles (EVs) act as key mediators of intercellular communication, but it remains unclear whether EV-encapsulated microRNAs (miRNAs) are involved in the crosstalk between endothelial cells (ECs) and monocytes in DKD. METHODS:EVs were isolated from monocytes and ECs under normal-glucose and high-glucose conditions. After characterization of EVs, differentially expressed miRNAs were screened. RNase/Triton functional assays, dual-luciferase reporter assays, gain- and loss-of-function experiments, and rescue experiments were further performed to validate the regulatory mechanisms associated with EV-miRNAs in vivo and in vitro. RESULTS:Under high-glucose conditions, monocyte-derived EVs aggravated EC injury by upregulating miR-191-3p, which targets CYLD and thereby activates the NF-κB pathway. Meanwhile, EC-derived EVs enhanced monocyte inflammation and adhesion by downregulating miR-615-3p, upregulating IFNGR2, and activating the STAT3 pathway. Targeted intervention of these key molecules effectively ameliorated cellular injury in vitro and partially relieved renal damage in vivo. CONCLUSIONS:This study confirms that EV-miRNA-mediated EC-monocyte crosstalk is involved in the pathogenesis of DKD, and establishes two core regulatory axes: miR-191-3p/CYLD/NF-κB and miR-615-3p/IFNGR2/STAT3. It also reveals an intrinsic link between microcirculatory dysfunction and immune injury in DKD, while the potential of these miRNAs as diagnostic biomarkers and therapeutic targets still remains to be further validated.

Attention deficit hyperactivity disorder (ADHD) is a prevalent neurodevelopmental disorder in children of school age. In mice with ADHD, glial fibrillary acidic protein (GFAP) levels were significantly higher, whereas glutamic acid decarboxylase 65 (GAD65) levels were lower. The goal of this study is to look at the potential function of GFAP and GAD65 antibodies (GAD65-Ab) in mice and children with ADHD by assessing GFAP and GAD65-Ab levels in the blood. An ADHD model was established in pregnant mice with 15 mg/kg of S-ketamine administration using intraperitoneal injection for 5 days from G14 to G18. Children were enrolled if they met in ADHD Rating Scale (ARS) in Diagnostic and Statistical Manual of Mental Disorders, 5th Edition (DSM-5) criteria for ADHD based on clinical interviews. Venous blood was obtained from children and mice under aseptic conditions and the serum was centrifuged and preserved. Clear supernatants from mice and children were processed simultaneously in duplicate, side by side, for ELISA assay with the calibrators supplied in the GFAP and GAD65-Ab kit. In mice with ADHD-like behaviors, there was a strong positive connection between serum GFAP levels and open-arm times in the elevated plus maze, which corresponded with the association between serum GFAP levels and hyperactivity-impulsivity ratings (r [Spearman] = 0.496, P = 0.004). The optimal cut-off value of serum GFAP levels as an indicator for auxiliary diagnosis of ADHD was projected to be 28.8 ng/ml based on the receiver operating characteristic (ROC) curve, yielding a sensitivity of 28.6

Background: Neuronal apoptosis is the core pathological mechanism of cerebral ischemic-reperfusion injury (CIRI); although Astragaloside IV (AS-IV) has demonstrated neuroprotective activity against CIRI, its specific molecular mechanisms underlying the regulation of this apoptosis-related pathway remain to be systematically elucidated. Methods: We establish an in vivo model of middle cerebral artery occlusion/reperfusion (MCAO/R) in rats and an in vitro model of oxygen-glucose deprivation/reperfusion (OGD/R) in PC12 cells. Six core apoptotic proteins, including CytC, Apaf-1, BAX, Bcl-2, Caspase3, and Caspase9, were detected using neurological function scoring, TTC/HE/Nissl staining, TUNEL staining, Western blot, and immunofluorescence techniques. Molecular docking and molecular dynamics simulation were utilized to analyze the binding affinity between AS-IV and the aforementioned apoptotic proteins. Results: Molecular docking and dynamics simulation demonstrated AS-IV stably binds six core apoptotic proteins, and comparative analysis with target-specific reference ligands identified Apaf-1 as its primary target with the most favorable binding properties. In rat MCAO/R models, AS-IV alleviated neurological deficits, reduced cerebral infarct volume and improved brain pathological damage; in PC12 cell OGD/R models, it decreased neuronal apoptosis. Western blot and immunofluorescence confirmed AS-IV downregulated pro-apoptotic proteins (cytoplasmic CytC, Apaf-1, BAX, cleaved-Caspase9/3) and upregulated anti-apoptotic Bcl-2. Conclusions: This study clarifies the anti-apoptotic molecular mechanism of AS-IV, it alleviates CIRI by targeting the CytC/Apaf-1 mitochondrial apoptotic pathway.

Background Cerebral ischemia‑reperfusion injury (CIRI) leads to poor stroke outcomes, partly due to sustained endothelial damage triggered by neutrophil extracellular traps (NETs) during the subacute phase. However, the exact downstream pathways through which NETs drive endothelial cell death, and whether Buyang Huanwu Decoction (BHD) intervenes in this process, remain unclear. Objective To investigate how NETs induce endothelial injury in CIRI and to evaluate the protective effects of BHD. Methods Chemical profiling of BHD aqueous extract and BHD‑containing serum was performed by UHPLC‑HRMS. A rat transient middle cerebral artery occlusion (tMCAO) model and an in vitro model of primary brain microvascular endothelial cells (BMECs) subjected to oxygen‑glucose deprivation/reperfusion (OGD/R) together with isolated NETs were used. Network pharmacology and transcriptomics were applied to predict key targets. NET formation was dynamically monitored. Functional experiments included a pyroptosis inhibitor, mitophagy modulators, BHD, and lentivirus‑mediated BNIP3 knockdown/overexpression. Results Absorbable components of BHD included calycosin‑7‑O‑β‑D‑glucoside, hydroxysafflor yellow A, and paeoniflorin. BHD improved neurological deficits, reduced infarct volume, and preserved blood‑brain barrier integrity in tMCAO rats. NETs peaked on day 3 after reperfusion and were suppressed by BHD. Mechanistically, NETs synergized with OGD/R to overactivate BNIP3, leading to dysregulated mitophagic flux and subsequent NLRP3/caspase‑1/GSDMD‑mediated endothelial pyroptosis. BNIP3 knockdown attenuated this cascade, whereas BNIP3 overexpression mimicked the damaging effects. BHD inhibited this NETs‑BNIP3‑pyroptosis cascade. In parallel, BHD partially restored mitochondrial dynamics (p‑DRP1, FIS1, MFN1) and promoted mitochondrial biogenesis (PGC‑1α, SIRT1, TFAM), consistent with its multi‑target regulation of mitochondrial quality control. Furthermore, under BNIP3 overexpression, BHD still reduced mitophagy and pyroptosis markers, indicating BNIP3‑independent protective effects. Conclusion These findings suggest that in CIRI, delayed NET formation overactivates BNIP3‑driven mitophagy and triggers endothelial pyroptosis. BHD exerts neurovascular protection by interfering with this NETs‑BNIP3‑pyroptosis axis while also improving mitochondrial dynamics and biogenesis as parallel protective mechanisms. This study provides experimental evidence for the pathogenic role of NETs and the multi‑target action of BHD in CIRI.

INTRODUCTION:The aim of this study was to investigate the mechanism of BZBS in the treatment of POF using network pharmacology and molecular docking, and to validate it through in vivo experiments. METHODS:Network pharmacology was used to construct chemical component-target pathway networks and protein-protein interaction networks to predict the potential targets of BZBS for the treatment of POF. The drug-target interactions were verified by molecular docking. Molecular dynamics simulations were performed to verify binding stability. Finally, experimental validation was performed. RESULTS:Network pharmacology analysis identified 220 BZBS compounds and 166 potential targets for POF treatment, with key core targets such as AKT1 and HIF-1α. Molecular docking showed that the main active ingredients of BZBS had a high affinity for POF, while molecular dynamics simulation verified stable interactions between AKT1 and Anhydroicaritin. It was confirmed that BZBS could ameliorate cisplatin CDDP-induced POF, regulate estrogen level, improve ovarian reserve, increase the expression of PI3K, AKT, HIF-1α, and VEGF proteins as well as immunofluorescence in ovarian tissues, and alleviate POF, which might be related to the activation of PI3K/AKT and HIF-1α/VEGF pathways. DISCUSSION:Through network pharmacology, molecular docking, molecular dynamics simulations, and in vivo experimental validation, it was preliminarily confirmed that BZBS ameliorates POF by activating PI3K/AKT and HIF-1α/VEGF pathways, which provides a theoretical basis for the subsequent studies on the treatment of POF with BZBS. However, our experiment lacks more in-depth clinical research. CONCLUSION:BZBS can improve symptoms of premature ovarian failure.

ETHNOPATHOLOGICAL RELEVANCE:BaZiBuShen (BZBS) is an innovative, patented traditional Chinese medicine known for its kidney-tonifying and anti-aging effects. It contains active ingredients such as flavonoids and amino acid analogues, which have anti-inflammatory and antioxidant properties, and is used to alleviate symptoms of "kidney essence" deficiency. AIM OF THE STUDY:This study evaluated the efficacy of BZBS on cyclophosphamide (CTX)-induced premature ovarian insufficiency (POI) and explored its possible mechanism of action. METHODS:POI models in rats were established using CTX to assess the therapeutic effects of BZBS. HPLC-MS was used to analyze the components, while ELISA was adopted to determine the serum hormone levels. Ovarian morphology and number of follicles were evaluated by H&E staining. The ultrastructure of mitochondria was examined by TEM. The expression levels of proteins related to ferroptosis and the NF2-YAP signalling pathway were analysed using immunohistochemistry, immunofluorescence, and Western blotting. RESULTS:In CTX-induced POI rats, BZBS treatment effectively restored ovarian weight, while simultaneously decreasing serum FSH and LH levels and increasing E2 levels. Histological analysis of the ovaries revealed that BZBS significantly increased the number of primordial, growing, and mature follicles, as well as reducing the number of atretic follicles. Furthermore, BZBS treatment mitigated ferroptosis by decreasing key markers Fe2+, TFR, ACSL4, and restoring the levels of GSH and GPX4. Additionally, BZBS modulated the expression of critical proteins involved in ferroptosis and cell signalling pathways. Specifically, it down-regulated p-RB1, while up-regulating SLC7A11 and RB1. Moreover, BZBS upregulates NF2 while downregulating YAP expression and its nuclear translocation, thereby regulating the NF2-YAP signaling pathway involved in ferroptosis. CONCLUSIONS:In a CTX-induced POI rat model, BZBS effectively restores hormonal levels, mitigates ovarian damage, and curbs excessive primordial follicle activation. It also modulates ferroptosis-related protein expression, activates the NF2-YAP pathway, and could provide a potential therapeutic approach for POI.

ETHNOPHARMACOLOGICAL RELEVANCE:Stroke is a common condition that poses a significant threat to human health. Buyang Huanwu Decoction (BYHWD) is a traditional treatment used for stroke management. However, the exact mechanism by which BYHWD mitigates cerebral ischemia-reperfusion by regulating calcium overload and restoring mitochondrial function is not yet fully understood. AIM:The objective of this research was to examine the neuroprotective properties of BYHWD in reducing the damage produced by cerebral ischemia/reperfusion (I/R) injury via the modulation of calcium overload and mitochondrial dynamics (MD). METHODS:MCAO/R model success was evaluated via PSI laser scatter flowmetry. The neurological function scores were assessed. The cerebral infarct (CI) volume was detected via TTC staining. NeuN expression was detected via immunohistochemistry, and degenerated neurons were observed via FJC staining. The mitochondrial permeability transition pore (mPTP), the mitochondrial membrane potential (MMP), and ATP were detected. The reactive oxygen species (ROS) content and the NAD+/NADH ratio were determined. The glutamate (Glu) and glutamine (Gln) contents as well as the Ca2+ concentration were determined. The expression of PKCε, p-PKCε, namely, Sirt5, GLS, Drp1, p-Drp1 616, Fis1, Opa1, and Mfn2 was determined via Western blotting. Immunohistochemistry was used to detect p-PKCε, which is expressed at high levels. Immunofluorescence was used to detect p-Drp1 616, Opa1 and Sirt5 fluorescence intensity. RESULTS:BYHWD treatment enhanced neurological function, decreased the amount of CI, mitigated neuronal damage, decreased mPTP opening, restored the MMP, increased ATP synthesis, and decreased the ROS content after brain I/R. It also increased PKCε, p-PKCε, Sirt5, GLS, Opa1 and Mfn2 expression; downregulated p-Drp1 616, Drp1 and Fis1 expression; elevated the NAD+/NADH ratio and Gln content; and decreased the Glu content and Ca2+ concentration. The effects of BYHWD were reversed by the administration of the PKCε inhibitor εV1-2. BYHWD administration led to increased PKCε mRNA expression. CONCLUSIONS:BYHWD modulates MD by diminishing calcium overload through the PKCε-Nampt-Sirt5 axis, which restores mitochondrial function and mitigates brain I/R damage.

BackgroundOxidative stress is widely acknowledged as a key pathogenic mechanism in diabetic nephropathy (DN). In recent years, the role of oxidative stress in DN has garnered increasing attention. However, no bibliometric analysis has yet been conducted on the relationship between oxidative stress and DN. This study aims to systematically analyze the relevant literature, identify trends in research, assess current hotspots, and predict future directions.MethodsWe retrieved literature related to oxidative stress and DN from the Web of Science Core Collection database. We analyzed data on publication volume, countries/regions, institutions, journals, keywords, and other relevant metrics using VOSviewer, the Bibliometrix R package, and CiteSpace.ResultsFrom 2014 to 2024, a total of 4076 publications related to oxidative stress and DN were published across 755 journals, showing a consistent upward trend each year. China and the United States are the leading contributors in this field and demonstrate close collaborative efforts. The top contributors by country, institution, journal, and author include: China (1919 publications), Jilin University and Central South University (69 publications each), BIOMEDICINE & PHARMACOTHERAPY (117 publications), and Prof. Sun Lin (33 publications). The most frequent keyword is “oxidative stress” (3683 occurrences). In the co-citation analysis, Alicic RZ’s 2017 study was the most cited (144 citations). These findings highlight the critical importance of investigating the pathogenesis of DN from the oxidative stress perspective.ConclusionThis study demonstrates a steady increase in research on oxidative stress in DN since 2014, highlighting its central role in the pathogenesis of DN. Future research should focus on the molecular mechanisms of oxidative stress in DN and explore its therapeutic potential, to provide new strategies for the prevention and treatment of DN.

Background/Objectives: Periostracum Cicadae (PC) is commonly used to treat chronic atrophic gastritis (CAG), but its underlying mechanisms are unclear. We investigated the therapeutic effects, active ingredients and molecular mechanisms of PC on CAG. Methods: We analyzed the components in the serum extract of PC by UHPLC-Q-Orbitrap-MS/MS. Then, we used rat and cell models to assess the impact of PC on CAG and employed network pharmacology and bioinformatics to predict key targets and active ingredients. Finally, we confirmed hub targets through experiments and molecular docking. Results: A total of 22 components were identified in the PC extract-containing serum using UHPLC-Q-Orbitrap MS/MS. Network pharmacology combined with molecular docking revealed that the protective effect was primarily mediated by three compounds: (Z)-akuammidine, chicoric acid, and columbianadin. And we revealed that c-Fos/c-Jun signaling pathways were crucial in therapy. PC extract-containing serum inhibited the vitality, migration, invasion, and multiplication of MC cells (model cells for CAG), induced apoptosis, and caused G0/G1 phase cell cycle arrest. The expression level of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), interleukin-1 beta (IL-1β) and gastrin 17 (G17) in the serum of CAG rats increased, while the expression level of pepsinogen I (PG I) and pepsinogen II (PG II) decreased. After 12 weeks of PC administration, these conditions were significantly improved. PC not only reduced the levels of antigen KI-67 (Ki67) and tumor protein p53 (P53) but also enhanced SRY-box Transcription Factor (SOX2). Simultaneously, PC down-regulated the expression of N-cadherin and Vimentin while up-regulating that of E-cadherin. Conclusions: PC inhibited epithelial–mesenchymal transition (EMT) via the c-Fos/c-Jun signaling pathway, thereby providing therapeutic benefits for CAG. Our study elucidates the mechanisms and material basis of PC in treating CAG, providing experimental evidence to support its clinical application.

Scutellaria baicalensis, a widely used medicinal herb in traditional Chinese medicine, is frequently employed in the treatment of diabetic nephropathy (DN). Its primary active constituent, baicalin, has shown significant therapeutic potential in animal models of DN; however, no comprehensive and systematic evaluation of its therapeutic effects and underlying mechanisms in DN has yet been conducted. This meta-analysis aimed to assess the efficacy of baicalin in DN treatment and elucidate its pharmacological mechanisms. Relevant studies were retrieved from databases including Web of Science, PubMed, Embase, CNKI, Wanfang Data, and VPCS, covering the literature up to November 2024. Study quality was evaluated using SYRCLE's risk of bias tool, and statistical analyses were performed with STATA 12. Primary outcomes included blood urea nitrogen (BUN), serum creatinine (SCR), and fasting blood glucose (FBG), while secondary outcomes encompassed urinary protein (UP), triglycerides (TG), total cholesterol (TC), inflammatory markers, fibrosis indicators, and oxidative stress parameters. Subgroup analyses, publication bias assessments, and sensitivity analyses were conducted to ensure result reliability. A total of 14 studies involving 221 rodents met the inclusion criteria. Baicalin significantly reduced BUN, SCR, FBG, TG, TC, UP, interleukin-6 (IL-6), interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), malondialdehyde (MDA), and fibronectin (FN) levels while enhancing superoxide dismutase (SOD) activity. These findings suggest that baicalin improves kidney function, reduces proteinuria, corrects lipid metabolism, and alleviates inflammation, oxidative stress, and fibrosis. This meta-analysis concludes that baicalin exhibits significant therapeutic potential in DN models, acting via anti-inflammatory, antioxidant, and antifibrotic mechanisms.

Cerebral ischemic-reperfusion injury (CIRI) involves mitochondrial dysfunction, with mitophagy playing a key role. Astragaloside IV (AS-IV) shows neuroprotective potential; however, its mechanisms related to mitochondrial function and apoptosis remain unclear. Methods: Using a rat MCAO/R model, we evaluated the AS-IV’s effects via neurological scores, TTC staining, and histopathology. Molecular assays and docking were used to analyze mitophagy (PINK1, Parkin, p62, ROS, Bcl-2, and BAX) and apoptosis markers. Results: AS-IV improved neurological function, reduced infarct volume, and alleviated neuronal/mitochondrial damage. It upregulated PINK1/Parkin, decreased p62, and modulated Bcl-2/Bax. Docking confirmed AS-IV binds PINK1/Parkin with high affinity. Conclusions: AS-IV protects against CIRI by regulating the PINK1/Parkin pathway, improving mitochondrial function, and inhibiting neuronal apoptosis, providing an experimental basis for the clinical use

Diabetic nephropathy (DN) is one of the most common and serious microvascular complications of diabetes mellitus (DM) and is the main cause of end-stage renal disease. Endothelial dysfunction caused by persistent hyperglycemia occurs at the initial stage of vascular disease. Moreover, persistent hyperglycemia is also a critical factor causing renal microcirculatory dysfunction. In recent years, many studies have confirmed that chronic hypoxia caused by microcirculatory dysfunction is one of the main mechanisms of kidney injury in patients with DM. Similarly, microcirculatory dysfunction damages renal tissue through interactions with other pathophysiological processes, thereby promoting the occurrence and development of DN. Thus, this article reviews the pathogenesis of renal microcirculatory dysfunction in DM and its interaction with stress, energy metabolism, and immunologic inflammation. Furthermore, a new idea was proposed to analyze the mechanism of kidney injury in DM from the perspective of microcirculatory dysfunction.

BACKGROUND:Attention deficit hyperactivity disorder (ADHD) is characterized by symptoms such as inattention, hyperactivity and impulsiveness, which significantly impact the healthy development of children. Our prior research demonstrated that exposure to S-Ketamine during pregnancy can lead to the development of ADHD, and existing studies have established a close association between astrocytes and the onset and progression of ADHD. The activation and inhibition of astrocytes are closely linked to neuropsychiatric dysfunction, and astrocytic NOD-like receptor protein 3 (NLRP3) has been reported to contribute to alterations in mental state and cognitive deficits. Thus, this study aims to investigate the role of astrocytes in ADHD by selectively modulating astrocyte function through Gq and Gi G protein-coupled receptors (GPCRs) and by specifically targeting the knockout of NLRP3. METHODS:Pregnant C57BL/6 J mice or mice with a specific deletion of NLRP3 in astrocytes were administered intraperitoneal injections of 15 mg/kg of S-ketamine for 5 consecutive days from gestational day 14 to 18 to establish an ADHD model. To modulate astrocyte activity in the hippocampal CA1 region, we administered astrocyte-specific Gq-Adeno-associated virus (AAV) or Gi-AAV into the CA1 and maintained treatment with CNO. At 21 days postnatally, we conducted open field test (OFT), novel object recognition (NOR), elevated plus maze (EPM) and fear conditioning (FC) in the offspring mice. Additionally, on postnatal day 21, we implanted electrodes in the CA1 region of the offspring mice for neurophysiological monitoring and investigated local field potentials (LFP) during NOR on postnatal day 27. Lastly, pathological assessments were conducted after euthanasia. RESULTS:Both the activation and inhibition of astrocytes in the hippocampal CA1 region improved impulsive-like behaviors and cognitive function in ADHD mice, reduced the power of theta (θ) oscillations during novel object exploration and decreased NLRP3-associated inflammatory factors, including cleaved caspase-1 and IL-18. Furthermore, compared to WT mice, astrocyte-specific NLRP3 conditional knockout mice demonstrated significantly reduced impulsive behavior and cognitive deficits, as well as a decrease in θ oscillation power and a reduction in NLRP3-associated inflammatory factors. CONCLUSIONS:Our data provide compelling evidence that the activation of astrocytes alleviated impulsive-like behaviors and cognitive dysfunction, possibly by reducing NLRP3-associated pyroptosis following changes in calcium levels within the astrocytes. The activation of astrocytes can be a potential therapeutic target for ADHD.

Background: Chronic atrophic gastritis precancerous lesions (PL-CAG) are characterized by the atrophy of gastric mucosal glands, often accompanied by intestinal metaplasia or dysplasia. Timely intervention and treatment can effectively reverse its malignant progression and prevent the onset of gastric cancer. Bombyx Batryticatus (BB) exhibits a range of pharmacological effects, including anticoagulation, antiepileptic properties, anticancer activity, and antibacterial effects. However, the pharmacological basis and mechanisms underlying BB’s efficacy in treating PL-CAG remain unclear. Methods: A three-factor modeling approach was implemented to develop a rat PL-CAG model, while the MNNG-induced PLGC (precancerous lesions of gastric cancer) cell model was served as a cell PL-CAG model. UPLC-QE-Orbitrap-MS/MS (Ultra performance liquid chromatography-quadrupole-electrostatic field orbital trap high-resolution mass spectrometry) was utilized to perform an in-depth analysis of the components in the plasma extract of BB. Leveraging network pharmacology, molecular docking analyses, and experimental validation, we initially elucidated the potential mechanisms through which BB mediates its therapeutic effects on PL-CAG at both in vivo and in vitro levels. Results: Prototype compounds of 42 blood-entering components were identified by UPLC-QE-Orbitrap-MS/MS analysis. Network pharmacology analysis and molecular docking studies indicate that the core targets are primarily enriched in the PI3K-Akt signaling pathway, and the key components, including Nepitrin, Quercetin 3-O-neohesperidoside, Rutin, and others, exhibited stable docking conformations with the first eleven pivotal targets. Both in vivo and in vitro experiments validated that BB may effectively treat PL-CAG via modulation of the PI3K-Akt signaling pathway. Conclusions: The therapeutic efficacy of BB in the management of PL-CAG may be achieved through the synergistic interaction of multiple components and targets, which may be more closely related to the inhibition of the PI3K/AKT signaling pathway. This approach will establish a solid experimental foundation and provide essential data for the clinical application of BB in treating PL-CAG, while also facilitating further research initiatives.

Mitochondria are an important organelle affecting the occurrence and development of ischemic stroke (IS). Although the role of mitochondria in IS has been paid attention to, the relevant biomarkers have not been identified, and the targeted treatment is still lacking. To further solve these problems, in this study, we combined and standardized GSE16561 and GSE58294 datasets in the GEO database as the test set, and GSE22255 as the training set. The mitochondria-related gene set was obtained from MitoCarta3.0 for study. R language was used to screen differentially expressed genes (DEGs), and weighted gene co-expression network analysis (WGCNA) was used to obtain the intersection genes. KEGG pathway enrichment and gene ontology (GO) analysis were performed. LASSO and Logistic methods were used to determine the diagnostic markers of mitochondrial-related IS. The correlation between it and the main immune cells and immune-related factors was analysed. The ROC curve was used to determine the diagnostic efficacy. The protein co-expression network, transcription factor, and miRNA prediction, and drug prediction by cMAP were performed against these markers. Molecular docking tested the binding energy. Transmission electron microscopy was used to observe the structure and morphology of mitochondria in cerebral microvascular endothelial cells of MCAO/R rats. Biochemical kits were used to detect the levels of MDA and SOD in blood and tissues. Elisa was used to detect the changes of the above targets in blood and tissues. In this study, a total of 38 intersection genes were obtained, and a diagnostic model composed of 3 genes (TST, SQOR, NRDC) was further established. KEGG and GO analysis showed that these genes were highly related to immunity and were involved in the immune activities related to neutrophils, CD4+T cells, CD8+T cells, and macrophages. In addition, a total of 42 proteins, 601 transcription factors, and 99 miRNAs related to TST, SQOR, and NRDC were predicted, and the interaction map was constructed. cMAP predicted 5 potential small molecule drugs and molecular docking suggested that W.13 had the best binding energy. In MCAO/R rats, the mitochondria in BMECs were severely damaged and fragmented, accompanied by a decrease in SOD activity and an increase in MDA level. In addition, the levels of TST and SQOR in blood and tissues were increased, and the levels of NRDC were decreased. Our study provides new insights into further understanding of IS from a mitochondrial perspective.

IntroductionLigustilide, a phthalide-derived bioactive compound abundantly found in traditional Chinese medicinal herbs such as Angelica sinensis (Danggui) and Ligusticum chuanxiong (Chuanxiong), has attracted increasing attention for its potential therapeutic benefits in ischemic stroke (IS). However, its clinical applications remain limited, and the comprehensive preclinical evidence regarding its efficacy and mechanisms of action is still unclear.Materials and methodsA systematic search of PubMed, Web of Science, and Embase was conducted to identify preclinical studies evaluating the effects of Ligustilide in IS animal models. A meta-analysis was performed to quantitatively assess the efficacy of Ligustilide in reducing infarct volume and improving neurological function. Additional analyses explored its potential mechanisms and the sources of heterogeneity.ResultsThe pooled results from 13 studies demonstrated that Ligustilide significantly reduced infarct volume (SMD = 3.26, 95% CI [2.31, 4.22], P < 0.05) and improved neurological scores (SMD = 1.64, 95% CI [1.13, 2.15], P < 0.05) in animal models of IS compared to control groups. Mechanistically, Ligustilide exerted protective effects by alleviating oxidative stress [lowering Malondialdehyde (MDA) levels (n = 3) and enhancing Superoxide Dismutase (SOD) (n = 2) and Glutathione (GSH) (n = 2) levels], suppressing inflammatory responses [reducing Tumor Necrosis Factor-alpha (TNF-α) (n = 3)], and a non-significant trend toward reduced apoptosis was also noted based on TUNEL staining (n = 2, P = 0.054), warranting further investigation. Sensitivity analyses confirmed the robustness of the findings. Subgroup analyses indicated that heterogeneity might be associated with differences in modeling methods, administration routes, and the use of multiple intervention doses.ConclusionThis systematic review and meta-analysis provides comprehensive preclinical evidence supporting the protective effects of Ligustilide in IS animal models through multi-target mechanisms. Future large-scale, high-quality animal studies and clinical trials are needed to further validate its therapeutic potential and facilitate its translational application.

Objectives: This study aims to elucidate the potential molecular mechanism of Sinomenine (SIN) in treating renal injury in Diabetic Nephropathy (DN) through network pharmacology, molecular docking, and in vivo validation. Materials and Methods: db/db mice were used as a DN model to evaluate the therapeutic effects of SIN on body weight, blood glucose levels, renal function, and histopathology. Network pharmacology and molecular docking were integrated to predict the potential molecular mechanisms of SIN in DN treatment. Subsequently, in vivo validation was performed on db/db mice using ELISA, Western blotting, RT-qPCR, immunofluorescence, and immunohistochemistry. Results: Firstly, we found that SIN (62.4 mg/kg) improved general conditions and renal function in db/db mice, alleviating renal pathological damage. Network pharmacology analysis identified IL-1β, IL-6, and TNF-α as key targets of SIN in DN. SIN reduced IL-1β, IL-6, and TNF-α levels by inhibiting the cGAS/STING signaling pathway and its downstream p-TBK1, p-IRF3, and NF-κB expression. Conclusions: SIN alleviates inflammatory injury in DN, potentially through the cGAS/STING pathway.

OBJECTIVE:To investigate the impact of Shenhua tablet (, SHT) on renal macrophage polarization and renal injury in mice with diabetic kidney disease (DKD)and to explore the potential mechanism involving the hypoxia-inducible factor-1α (HIF-1α) and pyruvate kinase M2 (PKM2) signaling pathway, along with the glycolysis metabolism pathway. METHODS:The animals were divided into the following groups: Model, Control, dapagliflozin, SHT low-dose, SHT medium-dose, and SHT high-dose. We assessed 24-hour urine protein (24 h-UTP) levels, urinary albumin-to-creatinine ratio, and regularly monitored fasting blood glucose during the treatment period. After treatment, we examined renal tissue structure, renal function (urea nitrogen, uric acid, creatinine, cystatin C, β2-microglobulin), and glycolysis in renal macrophages. Additionally, we observed macrophage polarization in renal tissue and measured inflammatory factors (tumor necrosis factor-α, interleukin-1β, interleukin-6, interleukin-10, monocyte chemoattractant protein-1) to assess the immunoinflammatory status of the renal tissue. Finally, we investigated the expression of the HIF-1α/ PKM2 signaling pathway in macrophages to explore its role in the glycolysis process. RESULTS:SHT shows a beneficial effect in treating DKD by reducing 24 h-UTP, regulating blood glucose levels, improving renal tissue structure, protecting renal function, inhibiting macrophage glycolysis, reducing macrophage transformation to the M1 state, and suppressing the expression of the HIF-1α/PKM2 signaling pathway. CONCLUSION:SHT may exert renoprotective effects by inhibiting macrophage glycolysis via the HIF-1α/PKM2 signaling pathway. This inhibition decreases macrophage M1 polarization and reduces immunoinflammatory injury in the renal tissue of DKD mice.